A typical cell cycle of a eukaryotic cell includes the M phase, which includes nuclear division (mitosis) and cytoplasmic division or cytokinesis and interphase, which begins with the G1 phase, proceeds into the S phase and ends with the G2 phase, which continues until mitosis begins, initiating the next M phase. In the S phase, DNA replication and histone synthesis occurs, while in the G1 and G2 phases, no net DNA synthesis occurs, although damaged DNA can be repaired. There are several key changes which occur during the cell cycle, including a critical point in the G1 phase called the restriction point or start, beyond which a cell is committed to completing the S, G2 and M phases.
Onset of the M phase appears to be regulated by a common mechanism in all eukaryotic cells. A key element of this mechanism is the protein kinase p34.sup.cdc2, whose activation requires changes in phosphorylation and interaction with proteins referred to as cyclins, which also have an ongoing role in the M phase after activation.
Cyclins are proteins that were discovered due to their intense synthesis following the fertilization of marine invertebrate eggs (Rosenthal, E. T. et al., Cell 20:487-494 (1980)). It was subsequently observed that the abundance of two types of cyclin, A and B, oscillated during the early cleavage divisions due to abrupt proteolytic degradation of the polypeptides at mitosis and thus, they derived their name (Evans, T. et al., Cell 33:389-396 (1983); Swenson, K. I. et al., Cell 47:867-870 (1986); Standart, N. et al., Dev. Biol. 124:248-258 (1987)).
Active rather than passive involvement of cyclins in regulation of cell division became apparent with the observation that a clam cyclin mRNA could cause activation of frog oocytes and entry of these cells into M phase (Swenson, K. I. et al., Cell 7:867-870 (1986)). Activation of frog oocytes is associated with elaboration of an M phase inducing factor known as MPF (Masui, Y. and C. L. Markert, J. Exp. Zool. 177:129-146 (1971); Smith, L. D. and R. E. Ecker, Dev. Biol. 25:232-247 (1971)). MPF is a protein kinase in which the catalytic subunit is the frog homolog of the cdc2 protein kinase (Dunphy, W. G. et al., Cell 54:423-431 (1988); Gautier, J. et al., Cell 54:433-439 (1988); Arion, D. et al., Cell 55:371-378 (1988)).
Three types of classes of cyclins have been identified to date: B, A and CLN cyclins. The B-type cyclin has been shown to act in mitosis by serving as an integral subunit of the cdc2 protein kinase (Booher, R. and D. Beach, EMBO J. 6:3441-3447 (1987); Draetta, G. et al., Cell 56:829-838 (1989); Labbe, J. C. et al., Cell 57:253-263 (1989); Labbe, J. C. et al., EMBO J. 8:3053-3058 (1989); Meier, L. et al., EMBO J. 8:2275-2282 (1989); Gautier, J. et al., Cell 60:487-494 (1990)). The A-type cyclin also independently associates with the cdc2 kinase, forming an enzyme that appears to act earlier in the division cycle than mitosis (Draetta, G. et al., Cell 56:829-838 (1989); Minshull, J. et al., EMBO J. 9:2865-2875 (1990); Giordano, A. et al., Cell 58:981-990 (1989); Pines, J. and T. Hunter, Nature 346:760-763 (1990)). The functional difference between these two classes of cyclins is not yet fully understood.
Cellular and molecular studies of cyclins in invertebrate and vertebrate embryos have been accompanied by genetic studies, particularly in ascomycete yeasts. In the fission yeast, the cdc13 gene encodes a B-type cyclin that acts in cooperation with cdc2 to regulate entry into mitosis (Booher, R. and D. Beach, EMBO J., 6:3441-3447 (1987); Booher, R. and D. Beach, EMBO J. 7:2321-2327 (1988); Hagan, I. et al., J. Cell Sci. 91:587-595 (1988); Solomon, M., Cell 54:738-740 (1988); Goebl, M. and B. Byers, Cell 54:433-439 (1988); Booher, R. N. et al., Cell 58:485-497 (1989)).
Genetic studies in both the budding yeast and fission yeast have revealed that cdc2 (or CDC28 in budding yeast) acts at two independent points in the cell cycle: mitosis and the so-called cell cycle "start" (Hartwell, L. H., J. Mol. Biol., 104:803-817 (1971); Nurse, P. and Y. Bissett, Nature 292:558-560 (1981); Piggot, J. R. et al., Nature 298:391-393 (1982); Reed, S. I. and C. Wittenberg, Proc. Nat. Acad. Sci. USA 87:5697-5701 (1990)).
In budding yeast, the start function of the CDC28 protein also requires association of the catalytic subunit of the protein kinase with ancillary proteins that are structurally related to A and B-type cyclins. This third class of cyclin has been called the C1n class, and three genes comprising a partially redundant gene family have been described (Nash, R. et al., EMBO J. 7:4335-4346 (1988); Hadwiger, J. A. et al., Proc. Natl. Acad. Sci. USA 86:6255-6259 (1989); Richardson, H. E. et al., Cell 59:1127-1133 (1989)). The CLN genes are essential for execution of start and in their absence, cells become arrested in the G1 phase of the cell cycle. The CLN1 and CLN2 transcripts oscillate in abundance through the cell cycle, but the CLN3 transcript does not. In addition, the CLN2 protein has been shown to oscillate in parallel with its mRNA (Nash, R. et al., EMBO J. 7:4335-4346 (1988); Cross, F. R., Mol. Cell. Biol. 8:4675-4684 (1988); Richardson, H. E. et al., Cell 59:1127-1133 (1988); Wittenberg, et al., 1990)).
Although the precise biochemical properties conferred on cdc2/CDC28 by association with different cyclins have not been fully elaborated, genetic studies of cyclin mutants clearly establishes that they confer "G1" and "G2" properties on the catalytic subunit (Booher, R. and D. Beach, EMBO J. 6:3441-3447 (1987); Nash, R. et al., EMBO J. 7:4335-4346 (1988); Richardson, H. E. et al., Cell 56:1127-1133 (1989)).
cdc2 and cyclins have been found not only in embryos and yeasts, but also in somatic human cells. The function of the cdc2/cyclin B enzyme appears to be the same in human cells as in other cell types (Riabowol, K. et al., Cell 57:393-401 (1989)). A human A type cyclin has also been found in association with cdc2. No CLN type cyclin has yet been described in mammalian cells. A better understanding of the elements involved in cell cycle regulation and of their interactions would contribute to a better understanding of cell replication and perhaps even alter or control the process.